Migration of fish leucocytes in vitro: the effect of factors which may be involved in mediating inflammation

Plaice (Pleuronectes platessa L.) neutrophils were isolated from the kidney on a discontinuous Percoll gradient and from the peritoneal cavity at the peak of a glycogen-elicited inflammatory response. The migratory ability of neutrophils was assessed using a 48-well microchemotaxis chamber, with an incubation of 1.5 h at 12 degrees C. The two neutrophil populations showed different responses to N-formylmethionyl-leucyl-phenylalanine (FMLP). Whereas kidney neutrophils only showed a significant enhancement of migration at 10(-7) M, inflammatory neutrophils exhibited a bimodal response, with one peak of migratory activity at 10(-9) M and a second at greater than 10(-6) M. Kidney neutrophils showed a consistent response with various concentrations of a 24 h culture supernatant of Vibrio alginolyticus. In every case increased migration was observed with 5-, 10- and 100-fold dilutions, with the latter two conditions producing a significant enhancement (p less than 0.01 and p less than 0.05 respectively). The undiluted and 2-fold diluted supernatant caused a decreased cell migration compared with control values. The supernatant from kidney neutrophils cultured with serum-opsonized, heat-killed V. alginolyticus produced greater migratory activity than neutrophils or the treated bacteria incubated alone (the controls). In each case, the enhanced activity of the supernatant was detectable by 1 h of incubation. By 4 h, the activity of the neutrophil/bacteria supernatant was significantly higher than that of the controls (p less than 0.01), but by 24 h had fallen to control levels. There was no evidence for a chemotactic response with FMLP, the bacterial supernatant or the neutrophil-derived factor and the responses were therefore assumed to be chemokinetic.